Shekar H. Shetty

Detection and identification of the blackeye cowpea mosaic strain of Bean common mosaic virus in seeds of cowpea from southern India

In different legume-growing regions of India, a total of 136 seed samples of cowpea (Vigna unguiculata (L.) Walp.) were collected and tested for the presence of the blackeye cowpea mosaic strain of Bean common mosaic virus, Cowpea aphid-borne mosaic virus, Cowpea mosaic virus, Cucumber mosaic virus and Bean yellow mosaic virus using growing-on test, enzyme linked immunosorbent assay, differential host test, electron microscopy and immuno-capture reverse transcription polymerase chain reaction. Among the 136 seedlots tested, 43 cowpea seedlots were found to be infected with BCMV-BlCM. The identity of three of these isolates as BCMV-BlCM was further supported by nucleotide sequencing in the 3′ region of the genome. The incidence of seeds carrying transmissible virus ranged from 0.67% to 13.49%. In most cases, only symptomatic seedlings in a growing-on test were found infected with the virus.

Management of Bean common mosaic virus strain blackeye cowpea mosaic (BCMV-BlCM) in cowpea using plant extracts

Abstract Efficacy of certain plant extracts in reducing Bean common mosaic potyvirus strain blackeye cowpea mosaic (BCMV-BlCM) disease in cowpea was evaluated. All the six botanicals Azadirachta indica, Boerhaavia diffusa, Bougainvillea spectabilis, Clerodendrum inerme, Psidium guajava, and Thuja occidentalis improved the germination and vigour index of cowpea. The disease incidence was reduced to 7% when 0.75% (w/v) of B. diffusa leaf extract was used as seed treatment under screen house conditions when compared to 80% in control. Under field conditions B. diffusa reduced the disease incidence up to 40% at 0.75% (w/v) concentration of extract. In spray treatment, B. diffusa and B. spectabilis reduced the disease incidence up to 13 and 12% under screenhouse conditions, whereas B. diffusa and C. inerme reduced the disease incidence up to 31 and 32% under field conditions. When plant extracts were mixed with BCMV-BlCM inoculum and young seedlings inoculated, B. spectabilis, C. inerme and M. jalapa extracts reduced the disease incidence up to 42, 40 and 48% respectively under screenhouse conditions when compared with the control.

Association between accumulation of allene oxide synthase activity and development of resistance against downy mildew disease of pearl millet

AbstractThe present study was aimed at understanding the possible association of allene oxide synthase (AOS), an enzyme implicated in the octadecanoid pathway during the pearl millet-downy mildew interaction. AOS 13-HPOT (13-hydroperoxy-9,11,15-octadecatrienoic acid) metabolizing activity assays assessed in various pearl millet cultivars with differential resistances against downy mildew revealed a positive correlation between cultivar resistance levels and AOS activities. Furthermore, the involvement of AOS in response to downy mildew was demonstrated by induction of AOS activity in both susceptible and resistant pearl millet cultivars during Sclerospora graminicola infection with higher induction observed in the resistant cultivar. Consistently, western blot analysis and tissue-blot immunoassay demonstrated the remarkable increase in AOS protein accumulation in the incompatible interaction. In addition, the tissue-blot immunoassay also showed the compartmentalization of AOS in the epidermis and vascular bundles of pearl millet seedlings. Expression analysis of a putative PgAOS1 gene revealed a marked difference in accumulation of PgAOS1 transcripts between contrasting plants, with pathogen-induced higher accumulation of the transcripts observed only in the resistant cultivar; a result which is in agreement with pathogen-induced AOS level and activity, indicating that PgAOS1 plays an important role in regulation of AOS level and activity in pearl millet upon S. graminicola infection. Our findings suggest an important role for AOS in regulation of responses to downy mildew disease in pearl millet. The differential AOS activities can potentially be used for selection of new disease-resistant pearl millet varieties, and the identified AOS-encoding gene(s) as genetic resource for development of enhanced downy mildew-resistant cultivars.

Serodiagnosis of pearl millet resistance to downy mildew by quantitating cell wall P/HRGP using polyclonal antiserum Pab-P/HRGP

Proline/hydroxyproline-rich glycoprotein (P/HRGP) level in pearl millet genotypes resistant to downy mildew increase after inoculation with the oomycete pathogen Sclerospora graminicola. Using purified P/HRGPs from pearl millet cell walls, polyclonal antibodies (Pab-P/HRGP) were raised in rabbit. Based on this antiserum, an enzyme immunoassay was developed that displays a linearity detection range from 0.01 to 10 μg P/HRGP. Western blot analysis, confirming the induction of three marker P/HRGPs in the infected resistant genotype, and immunocytochemical studies on P/HRGP localization either in epidermal peelings or in suspension-cultured cells demonstrated the specificity of the antiserum. Besides its characterization, Pab-P/HRGP was employed to screen various genotypes of pearl millet for fast, sensitive and specific detection of induced P/HRGPs upon infections. The results presented are discussed with presumed importance to downy mildew disease and the use of this new antiserum in pearl millet screening for disease resistance.

Genetic variation in Fusarium oxysporum f.sp. cubense isolates based on random amplified polymorphic DNA and intergenic spacer

Abstract Fusarium oxysporum f.sp. cubense (Foc) isolates were obtained from research centers and from the hot spot of fusarium wilt ‘Nanjangud’. The morphological features like colony appearance, growth rate, pathogenicity, optimum growth temperature and vegetative compatibility grouping (VCG) were determined. Molecular characteristics of these isolates were determined using random amplified polymorphism DNA (RAPD) analysis, and polymerase chain reaction – restriction fragment length polymorphism (PCR-RFLP) analysis of nuclear intergenic spacer rDNA (IGS). All the F. oxysporum f.sp. cubense isolates were pathogenic to banana cultivar ‘Nanjangud Rasabale’ but they did not induce any disease symptoms on cultivar ‘Cavendish’. The F. oxysporum isolate did not induce wilt symptoms on either ‘Nanjangud’ or ‘Cavendish’ cultivar. Based on RAPD data, 140 scoreable markers were obtained from 36 random decamer primers. Cluster analysis with Unweighted Pair-Group Method with Arithmetic mean (UPGMA) using genetic distance showed that the isolates belonged to three main groups. In general, isolate 1, 2, 4, 5 and 7 were in one group and isolate 3 and 6 were in another group. The grouping based on IGS data correlated with the grouping of isolates based on RAPD.